基于肿瘤微环境调控的功能转换型杂化多肽聚电解质刷载MDR1siRNA逆转结肠癌多药耐药性的递送机制研究

Low efficiency of gene delivery is thought to be the bottleneck in clinical gene therapy. Our research group has prepared and characterized a biodegrable peptide polyelectrolyte brushes (PEB). The ultra-low interface friction and strong osmotic pressure within the brushes brought about new physical properties for peptide PEB. Their unique features of peptide PEB had advantages in stability and stimulative responsibility compared with conventional polyelectrolyte gene carriers. This proposal based on our earlier studies, we design and evaluate the novel hybrid peptide PEB with tunable functions in tumor microenvironment to overcome inhibitory effects between long-circulating PEG and polycation of the carrier compositon in four key stages of in vivo delivery. The hybrid peptide PEB carrier will be optimized based on brush thickness, pKa and the grafting molar ratios to enhance the rates and extents of stimulative response to tumor microenvironment stimulation and achieve long circulation, high drug concentration at the tumor sites and highly effective gene silencing. The delivery and tunable function mechanisms of hybrid peptide PEB carried siRNA to down-regulate MDR1 mRNA in colorectal cancer will be specifically investigated. This novel hybrid peptide PEB carrier will provide a promising siRNA delivery system for reversal of multidrug resistance in human cancers which is leading to failure of chemotherapy.

给药系统不能满足基因药物体内有效递送是目前临床基因治疗的"瓶颈"。课题组已构建的可生物降解多肽聚电解质刷(Polyelectrolyte Brushes, PEB)具有超低摩擦系数和刷层高渗透压等特性，使多肽PEB的稳定性和环境刺激响应性显著强于普通聚电解质。本课题在前期研究基础上，针对基因载体设计中长循环材料PEG和基因复合材料阳离子聚电解质在体内四个递送关键环节存在相互制约减效的问题，构建肿瘤微环境调控的功能转换型杂化多肽PEB基因载体。通过刷层厚度、多肽pKa、接枝密度和杂化比例等方面优化，提高载体对肿瘤微环境响应速度和程度，达到长循环、肿瘤富集、高效基因沉默目的。探索杂化多肽PEB载MDR1siRNA逆转结肠癌多药耐药性的递送机制，明确递送功能转换的关键节点，实现体内有效递送。通过新型高效基因给药系统研发，本课题将为应用基因治疗解决多药耐药导致肿瘤化疗失败的临床难题提供一种新途径。

本课题针对基因递送系统中PEG 和阳离子聚合物存在相互制约减效的问题，依据肿瘤微环境与正常组织存在差异的特点，成功构建杂化多肽聚电解质刷（组氨酸杂化多肽聚电解质刷和谷氨酰胺杂化多肽聚电解质刷）。通过聚电解质pKa、接枝密度、聚电解质链摩尔比等方面优化，提高了组氨酸-谷氨酰胺功能转换型杂化多肽聚电解质刷基因载体对肿瘤微环境响应的速度和程度。实现在模拟正常生理条件下载体表面性质近中性，削弱和非靶细胞的相互作用，转染效率接近于0；而在模拟肿瘤微环境中载体表面发生功能转变，增强和靶细胞的亲和性，转染效率显著上升。应用杂化多肽聚电解质刷递送MDR1siRNA、SurvivinsiRNA和c-Myc siRNA多种功能小干扰RNA，在细胞和动物水平均显著提高了基因递送效率和基因沉默效率，进而有效逆转肿瘤多药耐药性和抑制了肿瘤生长。本课题为该基因递送系统的进一步应用提供实验和理论依据，为解决基因治疗中缺乏安全高效基因给药系统的临床难题和研发适合体内应用的基因递送系统具有科学意义。